Swabbing often fails to detect amphibian Chytridiomycosis under conditions of low infection load

PLoS One. 2014 Oct 21;9(10):e111091. doi: 10.1371/journal.pone.0111091. eCollection 2014.

Abstract

The pathogenic chytrid fungus, Batrachochytrium dendrobatidis (denoted Bd), causes large-scale epizootics in naïve amphibian populations. Intervention strategies to rapidly respond to Bd incursions require sensitive and accurate diagnostic methods. Chytridiomycosis usually is assessed by quantitative polymerase chain reaction (qPCR) amplification of amphibian skin swabs. Results based on this method, however, sometimes yield inconsistent results on infection status and inaccurate scores of infection intensity. In Asia and other regions where amphibians typically bear low Bd loads, swab results are least reliable. We developed a Bd-sampling method that collects zoospores released by infected subjects into an aquatic medium. Bd DNA is extracted by filters and amplified by nested PCR. Using laboratory colonies and field populations of Bombina orientalis, we compare results with those obtained on the same subjects by qPCR of DNA extracted from swabs. Many subjects, despite being diagnosed as Bd-negative by conventional methods, released Bd zoospores into collection containers and thus must be considered infected. Infection loads determined from filtered water were at least 1000 times higher than those estimated from swabs. Subjects significantly varied in infection load, as they intermittently released zoospores, over a 5-day period. Thus, the method might be used to compare the infectivity of individuals and study the periodicity of zoospore release. Sampling methods based on water filtration can dramatically increase the capacity to accurately diagnose chytridiomycosis and contribute to a better understanding of the interactions between Bd and its hosts.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Anura / genetics
  • Anura / microbiology*
  • Chytridiomycota / genetics
  • Chytridiomycota / isolation & purification*
  • Chytridiomycota / pathogenicity
  • DNA / genetics
  • DNA / isolation & purification
  • Infections / diagnosis*
  • Infections / genetics
  • Infections / microbiology
  • Polymerase Chain Reaction
  • Skin / microbiology
  • Skin / pathology

Substances

  • DNA

Grants and funding

The research was supported by the National Research Foundation of Korea (NRF), funded by the government of the Republic of Korea (MSIP) (grants 2010-0002767, 2012R1A1A2044449, and 2012K1A2B1A03000496 to BW), and by Seoul National University (Brain Fusion Program, Brain Korea 21 Program, and New Faculty Resettlement Fund grants to BW). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.